title: Formation of nucleoprotein filaments by mammalian DNA methyltransferase Dnmt3a in complex with regulator Dnmt3L.
abstract: The C-terminal domains of Dnmt3a and Dnmt3L form elongated heterotetramers (3L-3a-3a-3L). Analytical ultracentrifugation confirmed the Dnmt3a-C/3L-C complex exists as a 2:2 heterotetramer in solution. The 3a-3a interface is the DNA-binding site, while both interfaces are essential for AdoMet binding and catalytic activity. Hairpin bisulfite analysis shows correlated methylation of two CG sites in a distance of approximately 8-10 bp in the opposite DNA strands, which corresponds to the geometry of the two active sites in one Dnmt3a-C/3L-C tetramer. Correlated methylation was also observed for two CG sites at similar distances in the same DNA strand, which can be attributed to the binding of two tetramers next to each other. DNA-binding experiments show that Dnmt3a-C/3L-C complexes multimerize on the DNA. Scanning force microscopy demonstrates filament formation rather than binding of single tetramers and shows that protein-DNA filament formation leads to a 1.5-fold shortening of the DNA length.
authors.affiliation: Biochemistry Lab, School of Engineering and Science, Jacobs University Bremen, Bremen, Germany.
authors.forename: Renata Z
authors.initials: RZ
authors.lastname: Jurkowska
authors.forename: Nils
authors.initials: N
authors.lastname: Anspach
authors.forename: Claus
authors.initials: C
authors.lastname: Urbanke
authors.forename: Da
authors.initials: D
authors.lastname: Jia
authors.forename: Richard
authors.initials: R
authors.lastname: Reinhardt
authors.forename: Wolfgang
authors.initials: W
authors.lastname: Nellen
authors.forename: Xiaodong
authors.initials: X
authors.lastname: Cheng
authors.forename: Albert
authors.initials: A
authors.lastname: Jeltsch
mesh_terms: D000818:Animals; D004247:DNA / Q000378:metabolism / Q000648:ultrastructure; D004248:DNA (Cytosine-5-)-Methyltransferases / Q000737:chemistry* / Q000235:genetics / Q000378:metabolism; D019175:DNA Methylation; D000090205:DNA Methyltransferase 3A; D051379:Mice; D018625:Microscopy, Atomic Force; D009154:Mutation; D009698:Nucleoproteins / Q000648:ultrastructure*; D012436:S-Adenosylmethionine / Q000378:metabolism
chemical_list: C000715535:Dnmt3a protein, mouse; D009698:Nucleoproteins; D012436:S-Adenosylmethionine; D004247:DNA; C498796:Dnmt3l protein, mouse; D004248:DNA (Cytosine-5-)-Methyltransferases; D000090205:DNA Methyltransferase 3A